Fatty acid biosynthesis in higher plants has recently attracted increased interest because of the possible use of plant oils as renewable sources for reduced carbon. In plants, fatty acid biosynthesis occurs in the chloroplasts of green tissue or in the plastids of nonphotosynthetic tissues. The primary products in most plants are acyl carrier protein (ACP) esters of the saturated palmitic and stearic acids.
.DELTA..sup.9 stearoyl-acyl carrier protein desaturase (.DELTA..sup.9 desaturase) is a plastid localized non-membrane bound soluble desaturase that introduces the first double bond into saturated fatty acids (resulting in the corresponding mono-unsaturated fatty acids). Recently, several related soluble desaturases have been identified in the seed tissues of various plants that possess fatty acids with unusual double bond positions. Members of this class of soluble desaturases are specific for a particular substrate chain length and introduce the double bond between specific carbon atoms by counting from the carboxyl end of the fatty acid; for instance, the .DELTA..sup.9 desaturase is specific for stearoyl-ACP, and introduces a double bond between carbon 9 and 10. Initial desaturation reactions in animals and fungi, and subsequent desaturation reactions in plants, are mediated by a distinct class of fatty acid desaturases that are integral membrane proteins. Since most plants lack other desaturases that act on the 18:0 level, the ratio of saturated to unsaturated fatty acids in higher plants is mainly controlled by enzymes which catalyze the conversion of saturated to mono-unsaturated fatty acids. .DELTA..sup.9 desaturase cDNA encode precursor proteins containing an N-terminal transit peptide for targeting to the plastid. For safflower and castor, the 33 residue transit peptide is cleaved off to yield a 363 amino acid mature desaturase polypeptide with an apparent molecular weight of 37 kDa per subunit by SDS-PAGE. The enzyme occurs as dimers of approximately 70 kDa. The enzymatic reaction requires molecular oxygen, NAD(P)H, NAD(P)H ferredoxin oxido-reductase and ferredoxin.
Previous studies have shown that both soluble and membrane-bound .DELTA..sup.9 desaturases require non-haem iron for catalytic activity. More recently, spectroscopic analysis and amino acid sequence comparisons have established that the .DELTA..sup.9 desaturase contains a diiron cluster. This class of diiron proteins is characterized by two occurrences of the sequence motif E-X-X-H, spaced by approximately 100 amino acids, and includes the R2 subunit of ribonucleotide reductase and a methane monooxygenase hydroxylase component. A greater understanding of the catalytic mechanism of the acyl-ACP desaturase enzymes may enable the exploitation of such enzymes in the manufacture of plant seed oil.